Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.31 (beta-glucuronidase)
 7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The main purpose was to develop a high-performance liquid chromatography (HPLC)-based method to assay serotonin glucuronidation activity using liver microsomal fractions. Application of this method was then demonstrated by determining serotonin UDP-glucuronosyltransferase (UGT) enzyme kinetics using human liver microsomes and recombinant human UGT1A6. Interspecies differences were also evaluated using liver microsomes from 10 different mammalian species. 2. Incubation of liver microsomes with serotonin, UDP-glucuronic acid and magnesium resulted in the formation of a single product peak using HPLC with fluorescence and ultraviolet absorbance detection. This peak was confirmed as serotonin glucuronide based on sensitivity to beta-glucuronidase and by obtaining the expected mass of 352 with positive-ion mass spectrometry. 3. Following a preparative HPLC isolation, the structure of this metabolite was established as serotonin-5-O-glucuronide by (1)H-NMR spectroscopy. 4. Enzyme kinetic studies showed apparent K(m) and V(max) of 8.8 +/- 0.3 mM and 43.4 +/- 0.4 nmoles min(-1) mg(-1) protein, respectively, for human liver microsomes, and 5.9 +/- 0.2 mM and 15.8 +/- 0.2 nmoles min(-1) mg(-1), respectively, for recombinant UGT1A6. 5. The order of serotonin-UGT activities in animal liver microsomes was rat > mouse > human > cow > pig > horse > dog > rabbit > monkey > ferret. Cat livers showed no serotonin-UGT activity. Heterozygous and homozygous mutant Gunn rat livers had 40 and 13%, respectively, of the activity of the normal Wistar rat, indicating a significant contribution by a rat UGT1A isoform to serotonin glucuronidation. 6. This assay provides a novel sensitive and specific technique for the measurement of serotonin-UGT activity in vitro.
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PMID:Serotonin (5-hydroxytryptamine) glucuronidation in vitro: assay development, human liver microsome activities and species differences. 1262 59

Mitiglinide (MGN) is a new potassium channel antagonist for the treatment of type 2 diabetes mellitus. In the present study, a potential metabolic pathway of MGN, via carboxyl-linked glucuronic acid conjugation, was found. MGN carboxyl-glucuronide was isolated from a reaction mixture consisting of MGN and human liver microsomes fortified with UDP-glucuronic acid (UDPGA) and identified by a hydrolysis reaction with beta-glucuronidase and HPLC-MS/MS. Kinetic analysis indicated that MGN from four species had the highest affinity for the rabbit liver microsomal enzyme (K(m)=0.202 mM) and the lowest affinity for the dog liver microsomal enzyme (K(m)=1.164 mM). The metabolic activity (V(max)/K(m)) of MGN to the carboxyl-glucuronidation was in the following order: rabbit>dog>rat>human. With the assessment of MGN glucuronide formation across a panel of recombinant UDP-glucuronosyltransferase (UGT) isoforms (UGT1A3, UGT1A4, UGT1A6, UGT1A9, and UGT2B7), only UGT1A3 and UGT2B7 exhibited high MGN glucuronosyltransferase activity. The K(m) values of MGN glucuronidation in recombinant UGT1A3 and UGT2B7 microsomes were close to those in human liver microsomes. The formation of MGN glucuronidation by human liver microsomes was effectively inhibited by quercetin (substrate for UGT1A3) and diclofenac (substrate for UGT2B7), respectively. The MGN glucuronidation activities in 15 human liver microsomes were significantly correlated with quercetin (r(2)=0.806) and diclofenac glucuronidation activities (r(2)=0.704), respectively. These results demonstrate that UGT1A3 and UGT2B7 are catalytic enzymes in MGN carboxyl-glucuronidation in human liver.
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PMID:Carboxyl-glucuronidation of mitiglinide by human UDP-glucuronosyltransferases. 1735 41

Chloramphenicol (CP), a broad spectrum antibiotic, is eliminated in humans by glucuronidation. The primary UGT enzymes responsible for CP O-glucuronidation remain unidentified. We have previously identified the 3-O-CP (major) and 1-O-CP (minor) glucuronides by beta-glucuronidase hydrolysis, liquid chromatography-tandem mass spectrometry, and 1D/2D H NMR. Reaction phenotyping for the glucuronidation of CP with 12 expressed human liver UGT isoforms has identified UGT2B7 as having the highest activity for 3-O- and 1-O-CP glucuronidation with minor contributions from UGT1A6 and UGT1A9. The kinetics of CP 3-O-glucuronidation by pooled human liver microsomes (HLMs) exhibited biphasic Michaelis-Menten kinetics with the apparent high-affinity K(m1) and low-affinity K(m2) values of 46.0 and 1027 microM, whereas expressed UGT2B7 exhibited Michaelis-Menten kinetics with the apparent K(m) value of 109.1 microM. The formation of 1-O-CP glucuronide by pooled HLM and expressed UGT2B7 exhibited substrate inhibition kinetics with apparent K(m) values of 408.2 and 115.0 microM, respectively. Azidothymidine (AZT) and hyodeoxycholic acid (substrates of UGT2B7) inhibited 3-O- and 1-O-CP glucuronidation in pooled HLMs. In 10 donor HLM preparations, both CP 3-O- and CP 1-O-glucuronidation showed a significant correlation with AZT glucuronidation (UGT2B7) (r(s) = 0.85 and r(s) = 0.83, respectively) at 30 microM CP, whereas no significant correlation was observed between CP 3-O-glucuronidation and serotonin glucuronidation (UGT1A6) or propofol glucuronidation (UGT1A9) at this CP concentration. These results suggest that UGT2B7 is the primary human hepatic UDP-glucuronosyltransferase isoform catalyzing 3-O- and 1-O-CP glucuronidation with minor contributions from UGT1A6 and UGT1A9.
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PMID:Identification of human UGT2B7 as the major isoform involved in the O-glucuronidation of chloramphenicol. 2000 37